This proposal seeks to continue studies on ribozymes with non-cleavable internucleotide linkages and to exploit particular chemical modifications in internucleotide linkages in the study and development of new therapeutic agents that are aimed at specific target sequences in nucleic acids as a mechanism for the control of diseases. The modifications are "in-line" backbone changes in which the phosphodiester is converted to either a phosphono(5')methylene or phosphono(3')methylene linkage by substituting a methylene group for the 5'-oxygen or 3'-oxygen, respectively. This proposal is based on the following recent observations (1) The ADP and ATP analogues in which the 5'-oxygens are replaced with methylene groups serve as substrates for polynucleotide phosphorylase and phage T3 RNA polymerase, respectively. (2) Oligonucleotides containing all phosphono(5')methylene linkages appear to be isosteric with their natural counterparts in that they are able to participate in triple strand formation. (3) Because of the presence of the P-C bond, oligomers containing such linkages are completely resistant to the types of chemical and nuclease cleavage that involve breakage between the phosphorous and the 5'-oxygen in their natural counterparts. In-depth chemical, physical, enzymatic and biological studies will be performed on mononucleotides, oligonucleotide and nucleic acid analogues containing phosphonomethylene linkages. In addition substitution of a phosphono(5')methylene linkage at the cleavage point in a ribozyme substrate should permit formation of the ribozyme's active conformation without risk of its self-destruction during analysis. A second special application concerns the potential use of the mononucleotide analogues themselves as therapeutic drugs and it seems likely that one or more of the ribo- and deoxyribo-nucleoside triphosphate analogues containing a methylene group in place of the 5'-hydroxyl will be found to serve a substrates for one or more of the human polymerases (including reverse transcriptase).